PH indicator swabs for biomonitoring and diagnostics

ABSTRACT

Medical swabs incorporating a pH indicator in an absorbent crosslinked hydrogel matrix. The medical swabs can provide for visual monitoring of biological pH. The medical swabs can be used to monitor wounds, such as chronic wounds, burn wounds, surgical wounds, etc., as well as other biological fluids. The swabs can be used for early detection of chronic wounds and/or early detection of bacterial infections.

CROSS REFERENCE TO RELATED APPLICATION

This application claims filing benefit of U.S. Provisional PatentApplication Ser. No. 62/727,220, having a filing date of Sep. 5, 2018,entitled “pH-Indicating Swabs for Wound Biomonitoring and Diagnostics,”which is incorporated herein by reference for all purposes.

FEDERAL RESEARCH STATEMENT

This invention was made with Government support under Grant No. 1811949,awarded by the National Science Foundation, and under Grant No. R03EB026813, awarded by the National Institutes of Health. The Governmenthas certain rights in the invention.

BACKGROUND

The pH of biological fluids can provide a good determination of thestatus of an individual. For instance, the normal pH of intact skinranges from about 4.8 to about 6.0. When a wound occurs, the skin acidicmilieu and pH is disrupted, exposing the more neutral pH of theunderlying tissue (generally about pH 7.4). Over the course ofsuccessful healing and re-epithelialization the initial, more neutral,pH of the wound will drop due to various factors, including hypoxia andincreased production of lactic acid. An acidic pH environment isconsidered to be beneficial during healing by increasing fibroblastproliferation and migration, promoting epithelization, regulatingbacterial colonization, and facilitating the release of oxygen. Over thecourse of typical healing, an acute wound will pass through thephysiological stages of inflammation, tissue formation, and remodelingin a timely fashion if the wound is kept clean; the skin will return tothe healthy, slightly acidic range. If, however, wound healing isdelayed, the pH will oscillate and become increasingly alkaline overtime. At this stage, a wound can be considered to be a chronic wound.Chronic wound environments have been reported in the range of pH 7.15 to8.93.

Chronic wounds, such as leg ulcers, foot ulcers, and pressure sores ordecubitus ulcers caused by sustained external skin pressure, do not healnormally and can be detained in or not progress at all through thenormal wound healing stages. Chronic wounds require intensive levels ofcare, and even with intensive treatment, may take years to fully heal ormay never heal at all, causing long-term pain, as well as emotional andfinancial distress to the sufferer. An alkaline wound environment can beindicative of a chronic wound, and wounds of any sort having an elevatedalkaline pH have been shown to have lower rates of healing than woundsin which the pH is lower. Alkaline wounds are also more likely to becomeinfected and many wounds, especially chronic leg ulcers, are oftencolonized by intestinal, oral and resident dermal microorganisms.

The pH of biological fluids, with wound exudate being only one example,can affect many factors of the local environment, including oxygenrelease, angiogenesis, protease activity, macrophage and fibroblastactivity, and bacterial toxicity. High pH can be an indicator of anexisting infection, as pH increase from normal baseline is morefavorable for bioburden of pathogenic microorganisms. Moreover, somebacteria produce ammonia, which in itself is necrotizing, and which canimpair oxygenation of the tissues by further raising the pH and creatinga self-sustaining cycle.

It is known that the measurement of pH facilitates early detection ofinfection, which can enable early therapeutic intervention and improveoutcomes. Researchers have thus developed pH indicator compositions anddevices to monitor body fluids. For instance, U.S. Pat. Nos. 5,660,790;5,910,447; and 5,897,834, and U.S. Published Patent Application Nos.2009/0275071 and 2015/0308994 disclose devices and methods for analyzingthe pH of various bodily secretions. Commercial examples of pH sensitiveproducts include SwabCheck™ (Sigma), which includes a cellulose swabdesigned to collect a sample that is then transferred to an incubationtube; Amnicator™, which includes disposable swabs impregnated withnitrazine yellow dye; and VS-SENSE PRO™, which is a qualitative,visually readable swab for the evaluation vaginal fluid.

Unfortunately, many existing colorimetric pH indicator systems require areagent to extract the biological materials from the collection deviceand to develop a detectable color, and are often subject to dyeleaching, all of which adds to cost, complication, and potential error.Conventional pH indicators are also difficult to utilize as they requireregular calibration, include fragile electrodes that must remain wet,and lack flexibility often necessary to access tissue or fluids ofinterest.

In clinical practice, diagnosis of infection has been based on initialdetermination of secondary parameters such as odor, presence of localpain, heat, swelling, discharge, and redness. For instance, a wound maybe assessed visually, length and depth measurements may be taken, anddigital photography may be used to track the visual condition and sizeof a wound. Unfortunately, many of these clinical indicators have a lowpredictive value of infection and confirmation of the suspectedinfection is required before treatment can begin. Swabbing of fluid fromthe area of the suspected infection followed by microbiology testing isa standard option for confirmation of bacterial colonization andidentification of the strains associated with infection. Unfortunately,this process is time consuming and the time lag from onset of infectionto detection is one of the biggest drawbacks associated with existingclinical diagnostics, as delay in diagnosis can delay treatment andnegatively affect treatment outcome. Moreover, this process islabor-intensive and the multiple steps required in traditionalswab-based diagnoses increase the potential for error, such as samplingerrors, delays in transport of the swabs, errors in analyticalprocedures, and/or errors in reporting. While wound swabs have provenuseful, the wait for testing results of the sample and the potential forintroduction of error has limited their usefulness in the clinicalsetting, particularly for outpatients.

What is needed in the art is a convenient, rapid, and reliable method ofdetecting infection in early stages. There is an unmet need fordiagnostic reagents and methods that enable early diagnosis of clinicalinfection, and which permit clinical diagnosis prior to manifestation ofclinical symptoms of infection.

SUMMARY

According to one embodiment, disclosed is a medical swab that includes asurface and a crosslinked absorbent hydrogel matrix at the surface. Themedical swab can also include a pH indicator retained within thecrosslinked absorbent hydrogel matrix. The pH indicator exhibits avisually detectable color that is indicative of the pH of a fluid incontact with the pH indicator.

Also disclosed is a method for forming the medical swabs. For instance,a method can include forming a precursor solution comprising a pHindicator and a biocompatible polymer. The method can also includecrosslinking the biocompatible polymer to form a crosslinked absorbenthydrogel matrix with the pH indicator retained within the matrix.

A method for detecting the pH of a biological fluid is also disclosed,which can include contacting a biological fluid or a tissue that carriesor exudes the biological fluid with a medical swab, as described.Examination of the swab can visually determine the color of the pHindicator held in the swab, which can indicate the pH of the biologicalfluid. The method can be utilized in one particular embodiment fordetecting the pH of a wound and can facilitate early diagnosis ofinfection in the wound, e.g., in a chronic wound.

BRIEF DESCRIPTION OF THE FIGURES

A full and enabling disclosure of the present subject matter, includingthe best mode thereof to one of ordinary skill in the art, is set forthmore particularly in the remainder of the specification, includingreference to the accompanying figures in which:

FIG. 1 illustrates colorimetric transition of the anthocyanin swabs inbuffer solutions with different pH values. The original swab was purpleat pH 7.0 (center) and it changed color from red to pink in acidicsolutions (left) and blue to green (right) in basic solutions.

FIG. 2 illustrates colorimetric transition of litmus swabs in responseto buffer pH 4.8 and 9.2. Original litmus swab was purple at neutral pHof 7.0 (center), when exposed to acidic buffer turned red (left) and inbasic buffer turned blue (right).

FIG. 3 illustrates colorimetric transition of bromothymol blue swabs inresponse to buffer with different pH. Original swab was yellow (left),when exposed to neutral buffer turned light green (center) and in basicbuffer turned dark blue-green (right).

FIG. 4 illustrates colorimetric transition of the anthocyanin swabs inresponse to Pseudomonas aeruginosa and Staphylococcus aureus. Purpleoriginal swab (right) turned blue in the bacteria solutions.

FIG. 5 illustrates colorimetric transition of the litmus swabs inresponse to Pseudomonas aeruginosa. Purple original swab (right) turnedblue (left) in the bacteria solution.

FIG. 6 illustrates colorimetric transition of the bromothymol blue swabsin response to E. coli. Yellow original swab (right) turned blue (left)in the bacteria solution.

FIG. 7 illustrates the effect of swabs on cell viability of NHDFs. Cellviability (%) was calculated using an MTS assay after 24 h of exposure.No toxicity was observed.

Repeat use of reference characters in the present specification anddrawings is intended to represent the same or analogous features orelements of the present invention.

DETAILED DESCRIPTION

Reference will now be made in detail to various embodiments of thedisclosed subject matter, one or more examples of which are set forthbelow. Each embodiment is provided by way of explanation of the subjectmatter, not limitation thereof. In fact, it will be apparent to thoseskilled in the art that various modifications and variations may be madein the present disclosure without departing from the scope or spirit ofthe subject matter. For instance, features illustrated or described aspart of one embodiment, may be used in another embodiment to yield astill further embodiment.

The present disclosure is directed to medical swabs or swab-likestructures that permit direct contact with an area of a patient's body,for instance an area such as a wound that is suspected of harboring apathogenic micro-organism. The medical swabs incorporate a non-toxiccomposition that can provide for monitoring of biological pH in asimple, accurate, and rapid manner. Due to the ease of use and quick,visually detectable response of the medical swabs, pH monitoring can becarried out frequently and consistently, which can in one embodimentprovide an on-going and accurate understanding of the dynamic conditionof a biological fluid and the area from which the fluid is obtained.

The medical swabs include an absorptive crosslinked hydrogel and a pHindicator retained within the crosslinked hydrogel. The medical swabscan overcome disadvantages of previously known pH indicators byproviding a non-toxic composition that enables simple and rapidmonitoring of pH status of any area or biological fluid of interest, forinstance in on-going monitoring of a wound for potential chronic wounddevelopment or monitoring an area for local or systemic bacterialinfection. The swabs can be utilized in a one-step test, the results ofwhich can be interpreted by merely looking at the swab, with noinspection or color detection equipment or analysis necessary.

The pH indicator of a medical swab can be any suitable colorimetric pHindicator that is capable of exhibiting a relatively rapid color changein the visible spectrum (light having a wavelength of from about 380 nmto about 740 nm) with change in pH. A pH indicator can in one embodimentdetect pH between about pH 0 and about pH 14, though a pH indicator thatdetects pH within a smaller range is also encompassed. For instance, apH indicator may be utilized that detects pH between about pH 5 andabout pH 12, or about pH 6 and about pH 10 (e.g., a pH range expected inwound care). A suitable pH indicator can indicate changes in pH by wayof a color change along the visual spectrum, with visually identifiablecolors in the spectrum being associated with a particular pH. Inaddition, a pH indicator can exhibit a color change relatively rapidly,generally within a few minutes, e.g., about 5 minutes or less or about 1minute or less in some embodiments.

Examples of suitable pH indicators can include, without limitation,litmus, anthocyanin, nitrazine yellow, brilliant yellow, cresol red,bromcresol purple, chlorophenol red, bromothymol blue, thymol blue,bromoxylenol blue, neutral red, phenol red, xylenol blue, m-cresolpurple, orcein, erythrolitmin (or erythrolein), azolitmin, spaniolitmin,leucoorcein, and leucazolitmin, as well as combinations of indicators.

The medical swabs include the pH indicator retained within a crosslinkedhydrogel matrix. Retention of the pH indicator within the hydrogel canprevent color change due to unintended contact with a fluid other thanthe biological fluid of interest and can also prevent leaching of the pHindicator from the medical swab.

The hydrogel matrix of the medical swab can be an absorptive andnon-toxic material that can facilitate absorbing of a biological fluidof interest and movement of the fluid to the interior of the hydrogel,wherein it can contact the pH indicator retained therein. The hydrogelmatrix can be formed from one or more non-toxic, biocompatible polymersthat include, or can be modified to include, crosslinkablefunctionality. The matrix polymer can include synthetic and/or naturalhydrogel-forming polymers. By way of example, and without limitation,hydrogel matrix polymers can include alginates, collagen or derivativesthereof, cellulose or derivatives thereof, poly(lactic-co-glycolic acid)(PLGA) or derivatives thereof, polycaprolactone (PCL) or derivativesthereof, as well as combinations of different polymers, e.g., blends orcopolymers thereof.

According to one embodiment, the hydrogel matrix can include analginate. Alginate is a naturally occurring anionic biocompatiblepolymer with low toxicity composed of a variety of alginic acidsextracted from certain species of seaweeds. Alginate has beenextensively investigated for many biomedical applications, includinghighly absorbent wound dressings. Alginate contains blocks of(1,4)-linked β-D-mannuronate (M) and α-L-guluronate (G) residues.Alginate has strong hydrophilicity, and as such, can form a highlyabsorbent wound dressing. Alginates are commercially available as pads,ropes, or ribbons from a variety of different suppliers. Alginate can beattractive for incorporation in a hydrogel matrix in one embodiment, asit is known as an approved material for epidermal applications and hasexcellent biocompatibility and mechanical properties. Additionally,calcium alginate is known as a natural hemostat that can be removed froma wound site limited or no trauma and discomfort.

The hydrogel matrix polymer is not limited to alginates, however, andother non-toxic, biocompatible hydrogel-forming polymers can be utilizedin conjunction with or alternative to an alginate. For instance, in oneembodiment, a hydrogel matrix can incorporate collagen, or a derivativethereof, for instance in the form of gelatin. Collagen is one of themain protein components of bone, cartilage, tendons, ligaments, andskin. Gelatin can be obtained from collagen by acidic or basichydrolysis or thermal degradation of collagen, which leads to rupture ofthe collagen triple helix into the random coil structure of gelatin.

In one embodiment, the hydrogel matrix can include a cellulose polymeror a derivative of cellulose (e.g., cellulose acetate, sodiumcarboxymethyl cellulose, ethylcellulose, nitrocellulose, bacterialcellulose, etc.). Cellulose is the most abundant polysaccharide, and itis inexpensive with good processability, renewability, and ease ofphysical and chemical modification. It has good mechanical properties,good hydrolytic stability, low toxicity, and excellent biocompatibility.

PCL as may be incorporated in a wound dressing is a hydrophobic,semi-crystalline, resorbable, aliphatic polyester. The crystallinitydecreases with increasing molecular weight, and PCL exhibits goodsolubility and low melting point (59° C.-64° C.), as well as excellentblend-compatibility, making it attractive for application in disclosedwound dressings. PCL can be biodegradable, but the degradation andresorption kinetics of PCL are relatively slow due to its hydrophobicityand high crystallinity, and as such, it can be beneficial in someembodiments as a component of disclosed hydrogels. PCL can be blended orco-polymerized with other polymers, such as PLA or PLGA, in order tomodify its physical properties in a desirable fashion.

PLGA is a highly studied biodegradable polymer as may be incorporated indisclosed hydrogel. In vivo, it is hydrolyzed into the non-toxic lacticacid and glycolic acid monomers. PLGA is commercially available indifferent molecular weights and copolymer compositions. The rate ofbiodegradation of a PLGA polymer can be controlled through selection ofthe copolymer ratio and molecular weight.

To form the medical swab, an aqueous precursor solution can be formedthat can include the pH indicator, the hydrogel matrix polymer and anyother desired additives. For instance, a precursor solution can beformed including the pH indicator in an amount of from about 0.1% w/v toabout 2% w/v. Depending upon the nature of the pH indicator, it may bepreferred in some embodiments that the pH indicator is first dissolvedby use of a suitable non-aqueous solvent (e.g., an alcohol such asethanol or the like) and then combined with water to form an aqueouscomposition. Similarly, an aqueous solution including the pH indicatorcan include co-solvents, or the like, as are generally known in the art,to provide for dissolution of the pH indicator.

Following formation of an aqueous solution including the pH indicator,the hydrogel polymer can be added to the solution such that theprecursor solution can include from about 0.5% w/v to about 20% w/v ofthe polymer. A precursor solution can also include any useful additivesas are generally known in the art, which can be combined with the othercomponents of a precursor solution in any desired order and inquantities as would be evident to one of skill in the art.

By way of example, in one embodiment a precursor solution canincorporate a plasticizer, which can improve the mechanical propertiesand flexibility of the medical swabs. Examples of suitable plasticizerscan include, without limitation, dioctylphthalate; castor oil;diacetylated monoglycerides; diethyl phthalate; glycerin; mono- anddi-acetylated monoglycerides; polyethylene glycol; propylene glycol;triacetin; triethyl citrate; bis-(2-butoxyethyl) adipate; andbis-(2-ethylhexyl) sebacate polyvinyl alcohol, polyvinyl alcohol,glycerol, and polyethylene glycol. When included, a precursor solutioncan generally include a plasticizer component in an amount of from 0 toabout 20 w/v % of the precursor solution.

Other additives as may be included in a precursor solution can includematerials as known in the art. For instance, a medical swab canincorporate one or more biologically active agents such as antimicrobialagents, antiseptic agents, antifungal agents, etc. in the precursorsolution, which can then be retained in the crosslinked hydrogel matrix.Antimicrobial agents may include, for example, sources of oxygen and/oriodine (e.g., hydrogen peroxide or a source thereof and/or an iodidesalt such as potassium iodide); antimicrobial metals, metal ions andsalts, such as, for example, silver-containing antimicrobial agents(e.g., colloidal silver, silver oxide, silver nitrate, silverthiosulphate, silver sulphadiazine, or any combination thereof); or anycombination thereof.

Other examples of additives of a precursor solution can include, withoutlimitation, surfactants, colorants, chloride sources, and mixturesthereof. Additives to the precursor solution can encompass materialsthat are retained in the crosslinked hydrogel matrix of the medicalswab, as well as materials that are not retained in the final product.For instance, an additive, e.g., a surfactant, may serve a purposeduring formation of the medical swab and may be removed from the othercomponents following its intended use and during a later stage offormation of the medical swab.

In some embodiments, a precursor solution can include a crosslinkingagent and/or a crosslink initiator. In some embodiments, one or both ofa crosslinking agent and a crosslink initiator can contact the precursorsolution only at the time of crosslinking and following combination ofthe hydrogel polymer with the pH indicator to form the precursorsolution. Conventional biocompatible cross-linking agents as aresuitably used to provide the necessary mechanical stability and tocontrol the properties of a hydrogel can be included in a precursorsolution (or combined with a precursor solution at the time ofcrosslinking). When included in the precursor solution, the amount ofcrosslinking agent and/or crosslink initiator to be included will bereadily apparent to those skilled in the art. For instance, acrosslinking agent can be included in an amount of from about 0.01% w/vto about 0.5% w/v, from about 0.05% w/v to about 0.4% w/v, or from about0.08% to about 0.3% w/v, of the hydrogel precursor solution. Typicalcrosslinking agents can include, without limitation, tripropylene glycoldiacrylate, ethylene glycol dimethacrylate, triacrylate, and methylenebis acrylamide. In one embodiment, a cationic crosslinking agent can beutilized. For example, a polyvalent elemental cation such as Ca²⁺, Mg²⁺,Al³⁺, La³⁺, or Mn²⁺ can crosslink polymers of a hydrogel matrix.

When included, a precursor solution may include a crosslink initiator instandard amounts, e.g., up to about 5% w/v, for instance from about0.002% w/v to about 2% w/v. In one embodiment, a precursor solution caninclude a photoinitiator, such as, and without limitation to, benzoylradicals such as type I-α-hydroxy-ketones and benzilidimethyl-ketals(e.g., Irgacure 651, Irgacure 184, and Daracur 1173 as marketed by CibaChemicals), as well as combinations thereof.

A medical swab can include supporting substrates that can carry thecrosslinked hydrogel matrix. For instance, a medical swab can includethe crosslinked hydrogel matrix coated on an absorptive substrate thatcan facilitate absorption of a biological fluid of interest into thehydrogel matrix so as to encourage contact between the biological fluidand the pH indicator. An absorptive supporting substrate can be afibrous substrate, e.g., a woven or non-woven absorptive textile, or anabsorptive non-fibrous substrate, e.g., an open-pored absorptive foam.For instance, an absorptive supporting substrate can include a fibrouswadding, e.g., a cotton wool, or other cotton-based fibers, polyesterfibers, polyurethane fibers, etc.; a medical gauze, or the like; or anabsorbent foam, such as an open-celled hydrophilic polyurethane foam, asurgical sponge, or the like.

In such an embodiment, the precursor solution can generally be coated onthe supporting substrate prior to crosslinking. For instance, aprecursor solution can be coated on individual fibers of a fibroussupporting substrate and/or on the surface of a fibrous non-fibroussupporting substrate prior to crosslinking. Coating can be carried outby dip coating, spray coating, spin coating, or any other suitablecoating methodology.

In one embodiment, a medical swab can include the crosslinked hydrogelmatrix coated on a more stable, relatively non-pliable structuralsupport, for instance a rod or other holding device formed of, e.g., ahollow or solid structure of a molded plastic (e.g., polyester), paper,or wood. The hydrogel matrix can be applied directly on such astructural support or, alternatively, can be coated on an absorptivesupporting substrate, which in turn can be adhered to the structuralsupport. For instance, a medical swab can be in the form of a typicalcotton swab with one or both ends of a supporting rod carrying a cottonwool wadding. Individual fibers and/or the overall surface as a whole ofan end cotton wool segment of the swab can be coated with the precursorsolution prior to crosslinking.

It should be understood that the medical swab does not require anunderlying support substrate, and in some embodiments, the crosslinkedhydrogel matrix can exhibit suitable mechanical strength characteristicsto serve as a medical swab, without the need for any supportingsubstrates or structural supports.

Following formation of the precursor solution and any post-formationprocessing (e.g., coating, forming, etc.), the precursor solution can becrosslinked to form the crosslinked hydrogel matrix and to secure the pHindicator within the crosslinked hydrogel matrix. In some embodiments,crosslinking can be initiated upon contact of a crosslinking agent withthe precursor solution in conjunction with suitable crosslink initiationparameters as necessary (e.g., temperature or light). For instance, ahydrogel precursor solution containing sodium alginate as the hydrogelpolymer can be combined with a divalent cation (via e.g., a CaCl₂solution) and crosslinking can spontaneously occur via ion exchange. Inother embodiments, crosslinking can be initiated by providing suitablecrosslink parameters to initiate crosslinking of the precursor solutionwithout the necessity of contact with an externally suppliedcrosslinking agent.

A medical swab can be utilized to sample a biological fluid of anysource. For example, sample collection from the skin, mouth, nose,throat, sweat gland, urine, vagina or other locations, including bothinternal and external tissue locations, may be accomplished by bringingthe crosslinked hydrogel matrix that is impregnated with the pHindicator dye into contact with the body fluid of interest, for instanceby brushing or scraping a body surface with a swab. Upon contact, theabsorptive hydrogel matrix can absorb fluid at the surface. As the fluidis absorbed into the hydrogel matrix, it will contact the retained pHindicator, and within a short time, the color of the indicator willchange according to the pH value of the biological fluid, which in oneembodiment can indicate the presence, type, or concentration of bacteriapresent in the fluid or in the area from which the fluid was obtained.

In one particular embodiment, the medical swabs can be utilized inmonitoring the pH of wounds. Monitoring wound pH can help predict theprogression of wound healing, as the pH of a wound can be indicative ofthe natural biochemical processes of healing, and alkalinization of awound can play a part in chronic wound development. Furthermore,monitoring wound pH can provide early diagnosis of the presence ofbacteria in the wound and imminent or existing infection. Wound pH canalso impact the effectiveness of antibiotics or other treatments in awound bed, and recognition of pH modulation in a wound, as well asactive regulation of local pH, can be helpful in guiding managementpractices and in determining effective treatment strategies, which canimprove patient outcomes.

Any type of wound can be monitored by use of the medical swabs, e.g.,acute wounds, chronic wounds, burns of any degree, internal wounds,external wounds, etc. For instance, wounds including ulcers, burns,sunburns, traumatic injuries, bedsores, diabetic wounds, surgicalwounds, and the like can be monitored. The medical swabs are not limitedto wound treatment, however, and can be beneficially utilized intreatment of any form of skin disease or irregularity as well as for usein monitoring other biological fluids and tissue types. Non-limitingexamples of biological fluids and tissues as may be monitored for pHabnormality by use of the medical swabs can include, without limitation,wounds and wound exudate, sweat or other skin exudate, vaginal surfacesand vaginal secretions, urine, saliva, amniotic fluids, blood, mucus,and nasal surfaces. Alterations in pH as may be detected by use of themedical swabs can provide for early diagnosis of serious medicalconditions, in addition to pathogenic infections including ischemicsyndromes such as coronary or peripheral arterial disease andangiogenesis-dependent disease.

Beneficially, the medical swabs can provide a qualitative,visually-readable result that is easily accessible to the clinicians, aswell as non-clinical individuals, who wish to evaluate wound status. Insome embodiments, the swabs can be used in conjunction with other, moretraditional clinical examination. For instance, following determinationof irregular pH in a fluid or tissue, further examination of the areacan be carried out, such as more traditional diagnostic approaches knownfor determination of the number(s) and type(s) of pathogenic flora atthe site using traditional laboratory and/or clinical diagnosticprocedures. However, due to the objective measurement of pH state of abiological fluid available by assessing the color of the swab aftercontact of the swab with the area of interest, a treatment plan can beinitiated prior to further examination by more traditional approaches,and the treatment plan can be based on the status of the wound asdetermined by the color of the swab applied to the surface. Earlierinitiation of treatment can lead to improved outcomes for patients, aswell as shorter treatment periods and decreased treatment costs.

The present disclosure may be better understood with reference to theExamples set forth below.

Example

Preparation of Litmus, Anthocyanins, and Bromothymol Blue

Anthocyanins solution was prepared by adding 55 g of chopped red cabbageto 90 mL of DI water, keeping at 90° C. for 1 hour, and filtering toobtain 90 mL of the final solution. About 0.1 g litmus dye (TCI, Tokyo,Japan) was added in 15 mL DI water and stirred to complete dilution.Bromothymol blue solution (Sigma Aldrich, USA) (0.1% w/v) was preparedby adding the dye to the water containing 0.2% v/v ethanol.

Preparation of Alginate Swabs Incorporated with pH Sensitive Dye

A solution of sodium alginate/glycerol (2% w/v-10% w/v)(Sigma-Aldrich-Fisher Scientific) was prepared in anthocyanin solutionand homogenized at 800 rpm using stirrer for 1 hour. Homogenizedsolution was put into a sonication bath to remove the trapped airbubbles. Additionally, alginate/glycerol (2% w/v-10% w/v) containing0.5% w/v litmus solution was prepared and homogenized for 1 hour. Asolution of sodium alginate/bromothymol blue (2% w/v-0.1% w/v) wasprepared by adding alginate to the dye solution following stirring for 1hour.

Cotton swabs were coated through dipping in the alginate solutionincorporated with a pH indicator dye solution for 1 hour followed bydipping in 2% w/v CaCl₂ for 10 minutes as crosslinker. pH indicatorswabs were washed with DI water for 5 minutes to remove unreacted andexcess CaCl₂ and dye.

Analysis of pH-Sensitive Swabs in Response to Different pH Environments

A glass microelectrode pH meter (VWR, USA) was used to measure the pH ofthe solutions of 0.1M potassium phosphate buffer, pH ranging from 2.4 to12.0. The indicator swabs were exposed to the buffer solution, andafterwards, the color was photographed.

Colorimetric Detection of Bacterial Infections

A strain of Gram-negative Pseudomonas aeruginosa and Gram-positiveStaphylococcus aureus and E. coli were cultured in Tryptic soy broth.These strains are highly prevalent in wounds and infections caused bythese pathogens remain a common complication in acute and chronicwounds. Following 24-hour incubation, 200 μL bacteria in culture wasdeposited on the pH indicator swabs and photographed.

Cytotoxicity Assay (MTS)

In order to determine cell viability, normal human dermal fibroblasts(NHDFs) were cultured in growth media DMEM/10% FBS to reach 80%confluency. Cells were seeded at a density of 1×10⁵ cell per 6 wellplate in a total volume of 2 mL in each well. Cells were incubated at37° C. and 5% CO₂ to allow for cell attachment and reach confluency. pHindicator swabs with 1 cm plastic support stick were sterilized throughUV radiation for 2 hours and were soaked in 2 mL growth media for 2hours. Eluates from the test swabs were added to the cells and exposedfor 24 hours; untreated cells were used as control. Following 24-hourincubation, media containing 20% MTS solution was replaced with eluatemedia and incubated for 2 hours. Colored formazan absorbance which wasproduced by bio reduction of MTS tetrazolium compound in live cells wasread at 490 nm using a Spectramax 190 spectrophotometer and cellviability was assessed.

Results

FIG. 1 demonstrates a gradient of colors for anthocyanin loaded swabswhen exposed to buffer with different pH 2.4, 4.84, 7.0, 9.2, and 12.0.The original swab had pH of 7.0. Anthocyanin impregnated swabs turnedred, pink, purple, blue, and green, respectively when exposed to therespective pH. Anthocyanins are a class of halochromatic substanceswhich respond to the level of acidity in an environment with a colorchange. Anthocyanins pigments are responsible for the red, purple, andblue colors of many fruits and vegetables, including red and purplecabbage, beets, purple carrots, elderberry, hibiscus, purple potato, redand black currants, blueberries, cherries, purple grapes, and red andblack raspberries. Anthocyanin pigment has a color gradient from orangeto blue in the visible light spectrum, following the range of possiblepH values.

FIG. 2 demonstrates litmus dye incorporated indicator swab exposed todifferent pH buffer solution. The litmus impregnated swab was originallyred under acidic conditions, pH 4.2, turned blue under basic conditions,pH 9.2, and was purple in neutral pH 7.0.

FIG. 3 demonstrates bromothymol blue loaded swab color change inresponse to pH changes. The swab was originally yellow, and swabschanged color to light green at pH of 7, and then shifted to blue-greenat the higher pH of 8.

FIG. 4 demonstrates color change of anthocyanin loaded swabs exposed to200 μL of bacteria strains. The original purple color changed to blue in10 minutes for both bacteria strains.

FIG. 5 demonstrates litmus dye loaded swabs exposed to 200 μL ofbacteria strains. The original purple swab turned blue in 10-minuteexposure to bacteria in culture.

FIG. 6 demonstrates bromothymol blue loaded swab exposed to 200 μL of E.coli bacteria strain. The color of the swab changed in 1 minute uponexposure to bacteria in culture.

FIG. 7 demonstrates cell viability results following 24-hour treatmentwith the swab extract. No cell toxicity was observed after 24 hours ofexposure of swab extract to fibroblast cell line.

While certain embodiments of the disclosed subject matter have beendescribed using specific terms, such description is for illustrativepurposes only, and it is to be understood that changes and variationsmay be made without departing from the spirit or scope of the subjectmatter.

What is claimed is:
 1. A medical swab comprising: a swab surface; acrosslinked absorbent hydrogel matrix comprising a biocompatible polymerat the swab surface; a photoinitiator; and a pH indicator retainedwithin the crosslinked absorbent hydrogel matrix, the pH indicatorexhibiting a detectable color change in the visible spectrum that isindicative of the pH of a fluid in contact with the pH indicator.
 2. Themedical swab of claim 1, the crosslinked absorbent hydrogel matrixcomprising a plasticizer.
 3. The medical swab of claim 2, theplasticizer comprising dioctylphthalate, castor oil, diacetylatedmonoglycerides, diethyl phthalate, glycerin, mono- and di-acetylatedmonoglycerides, polyethylene glycol, propylene glycol, triacetin,triethyl citrate, bis-(2-butoxyethyl) adipate, and bis-(2-ethylhexyl)sebacate polyvinyl alcohol, polyvinyl alcohol, glycerol, or polyethyleneglycol.
 4. The medical swab of claim 1, the biocompatible polymercomprising an alginate, a collagen, a gelatin, a cellulose, apoly(lactic-co-glycolic acid), a polycaprolactone, or derivatives orcombinations thereof.
 5. The medical swab of claim 1, the pH indicatorcomprising litmus, anthocyanin, nitrazine yellow, brilliant yellow,cresol red, bromcresol purple, chlorophenol red, bromothymol blue,thymol blue, bromoxylenol blue, neutral red, phenol red, xylenol blue,m-cresol purple, orcein, erythrolitmin (or erythrolein), azolitmin,spaniolitmin, leucoorcein, leucazolitmin, or any combination thereof. 6.The medical swab of claim 1, the medical swab further comprising asupport substrate, wherein the crosslinked absorbent hydrogel matrix iscoated on the support substrate.
 7. The medical swab of claim 6, thesupport substrate comprising an absorptive substrate.
 8. The medicalswab of claim 7, the absorptive substrate comprising fibers, a foam, ora sponge.
 9. The medical swab of claim 6, the support substratecomprising polyurethane, polyester, or cotton.
 10. The medical swab ofclaim 6, the support substrate comprising a hollow or solid structure ofmolded plastic, paper or wood.
 11. The medical swab of claim 1, whereinthe photoinitiator comprises a benzoyl radical.